title: Mutation of the IFNAR-1 receptor binding site of human IFN-alpha2 generates type I IFN competitive antagonists.
abstract: Type I interferons (IFNs) are multifunctional cytokines that activate cellular responses by binding a common receptor consisting of two subunits, IFNAR-1 and IFNAR-2. Although the binding of IFNs to IFNAR-2 is well characterized, the binding to the lower affinity IFNAR-1 remains less well understood. Previous reports identified a region of human IFN-alpha2 on the B and C helices ("site 1A": N65, L80, Y85, Y89) that plays a key role in binding IFNAR-1 and contributes strongly to differential activation by various type I IFNs. The current studies demonstrate that residues on the D helix are also involved in IFNAR-1 binding. In particular, residue 120 (Arg in IFN-alpha2; Lys in IFN-alpha2/alpha1) appears to be a "hot-spot" residue: substitution by alanine significantly decreased biological activity, and the charge-reversal mutation of residue 120 to Glu caused drastic loss of antiviral and antiproliferative activity for both IFN-alpha2 and IFN-alpha2/alpha1. Mutations in residues of helix D maintained their affinity for IFNAR-2 but had decreased affinity for IFNAR-1. Single-site or multiple-site mutants in the IFNAR-1 binding site that had little or no detectable in vitro biological activity were capable of blocking in vitro antiviral and antiproliferative activity of native IFN-alpha2; i.e., they are type I IFN antagonists. These prototype IFN antagonists can be developed further for possible therapeutic use in systemic lupus erythematosus, and analogous molecules can be designed for use in animal models.
authors.affiliation: Department of Molecular Genetics, Microbiology, and Immunology, UMDNJRobert Wood Johnson Medical School, Piscataway, New Jersey 08854, USA.
authors.forename: Manjing
authors.initials: M
authors.lastname: Pan
authors.forename: Eyal
authors.initials: E
authors.lastname: Kalie
authors.forename: Brian J
authors.initials: BJ
authors.lastname: Scaglione
authors.forename: Elizabeth S
authors.initials: ES
authors.lastname: Raveche
authors.forename: Gideon
authors.initials: G
authors.lastname: Schreiber
authors.forename: Jerome A
authors.initials: JA
authors.lastname: Langer
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D001665:Binding Sites / Q000235:genetics; D002417:Cattle; D004195:Disease Models, Animal; D006801:Humans; D016898:Interferon-alpha / Q000037:antagonists & inhibitors* / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D008180:Lupus Erythematosus, Systemic / Q000188:drug therapy / Q000235:genetics / Q000378:metabolism; D051379:Mice; D011485:Protein Binding / Q000235:genetics; D017433:Protein Structure, Secondary / Q000235:genetics; D053633:Receptor, Interferon alpha-beta / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism
chemical_list: C507964:IFNAR1 protein, human; C508033:IFNAR2 protein, human; C508032:Ifnar1 protein, mouse; C508034:Ifnar2 protein, mouse; D016898:Interferon-alpha; D053633:Receptor, Interferon alpha-beta